The goal of Kevin A. Keane, DVM is to become a faculty member at an institution conducting research on viral diseases. This training will occur via mentored investigation and coursework in fulfillment of his Ph.D. requirements for the Department of Pathology at Colorado State University. Dr. James C. DeMartini will serve as the mentor. The applicant's training will employ an animal model to investigate lentivirus-induced pulmonary disease. The applicant will also complete coursework in basic sciences and participate in other scholarly activities to become a biomedical scientist. The proposed research will investigate the effect of ovine lentivirus (OvLV) infection of alveolar macrophages in the pathogenesis of lymphoid interstitial pneumonia (LIP) as a model of this "non-AIDS" manifestation of HIV infection. The hypothesis is that dysregulation of alveolar macrophage homeostatic mechanisms occurs upon ovine lentivirus infection. This dysregulation is manifested through aberrant cytokine production and altered cell surface molecule expression that exaggerate T cell proliferation within the lungs in response to viral antigen and non-specific stimuli. Specific aim 1 is to correlate the sequential development of pulmonary lymphocyte proliferation in ovine lentivirus infected lambs with localization and phenotype of virus-infected pulmonary macrophages. Bronchoalveolar lavage cells and lung tissue will be stained by immunohistochemistry for macrophage activation molecules and lymphocyte accumulation (CD1, CD4, CD8, MHC II, B7, and CD25). Correlations with macrophage infection rates (in situ hybridization) and lesion severity will be calculated. Specific aim 2 is to examine the in vivo functional changes associated with cytokine from OvLV-infected alveolar macrophages and correlate with lymphocyte proliferation rates. RT-PCR will be used to measure mRNA for the TNF-a, IL-1B, IL-6, IL-10, IL-12 and INOS. Proliferation rates will be quantified using immunohistochemistry for BrDU, Ki67 and PCNA. Specific aim 3 is to examine in vitro functional changes in OvLV-infected alveolar macrophages and their effect on lymphocyte proliferation rates in response to mitogen. Cytokine mRNA from these cultures will be measured by RT-PCR similar to aim 2. Nitric oxide will be quantified by the Griess assay. Similar assays will be performed with nitric oxide production blocked by N-methyl-L-arginine. Data will be analyzed using digital image analysis. Conclusions derived from these studies will provide insight into the role of virus infected alveolar macrophages in the initiation and perpetuation of pulmonary T cell proliferation and will contribute to the knowledge of how alveolar macrophages maintain immune homeostasis in the lung.